A major problem in chemical countermeasure discovery is the potential emergence of novel agents for which there are no known antidotes or post exposure therapies. Classic countermeasures have often been discovered only serendipitously or have taken years to develop. As novel chemical threats emerge (in the form of novel chemical warfare agents or environmental pollutants), the speed with which we are able to understand and counteract each threat will determine the magnitude of its societal impact. One promising approach for rapidly identifying and manipulating the molecular pathways underlying the response to any chemical threat is the use of phenotype-based chemical screens. Models can be developed in which specific toxicants result in reproducible phenotypes in cells or whole organisms. By subjecting these models to high- throughput screening (HTS), small molecules could be identified that reverse the phenotype through a variety of novel mechanisms. Small molecules discovered by these screens would be excellent lead compounds for novel countermeasures and powerful tools for dissecting toxicity pathways, which may in turn point to additional countermeasure targets. In this application, we outline a process for rapid development of in vivo organophosphate toxicity models and their use in discovery of novel organophosphate countermeasures. Although we expect the proposed project will lead directly to development of novel organophosphate countermeasures, we also expect it to serve as a model for rapid countermeasure development that can be applied broadly to other existing and emerging chemical threats. Specifically, we propose: Aim 1. To develop and validate in vivo assays for organophosphate toxicity in the zebrafish. We will seek to identify physiological responses that are surrogates for known human responses to organophosphates and that can be scaled for high throughput in vivo screening. Aim 2. To scale these novel assays for automated high-throughput screening in multiwell plates. We will develop assays for organophosphate toxicity that can be performed automatically in 96-well format. Aim 3. To identify novel compounds that counteract the effects of prior organophosphate exposure. High-throughput screening will identify compounds that facilitate recovery from organophosphate exposure